High Stromal SFRP2 Expression in Urothelial Carcinoma Confers an Unfavorable Prognosis

Abstract

Background: Urothelial carcinoma (UC) patients often bear clinical and genetic heterogeneity, which may differ in management and prognosis. Especially, patients with advanced/metastatic UC generally have a poor prognosis and survive for only few months. The Wnt/β-catenin signaling is found to be highly activated in several cancers, including UC. However, accumulated evidence has shown discordance between the Wnt/β-catenin signaling and UC carcinogenesis. Accordingly, we aim to get a better understanding of the molecular characterization of UC, focusing on the Wnt signaling, which may add value to guiding management more precisely.

Patients and methods: Clinical data and pathological features were retrospectively surveyed. The correlations of secreted Frizzled-related protein 2 (SFRP2) immunoexpression with clinicopathological features were analyzed by Pearson's chi-square test. The Kaplan-Meier method with a log-rank test was employed to plot survival curves. All significant features from the univariate analysis were incorporated into the Cox regression model for multivariate analysis.

Results: Following data mining on a transcriptome dataset (GSE31684), we identified that 8 transcripts in relation to the Wnt signaling pathway (GO: 0016055) were significantly upregulated in advanced/metastatic bladder tumors. Among these transcripts, the SFRP2 level showed the most significant upregulation. Additionally, as SFRP2 is a putative Wnt inhibitor and may be expressed by stroma, we were interested in examining the immunoexpression and clinical relevance of stromal and tumoral SFRP2 in our urothelial carcinoma cohorts containing 295 urinary bladder UC (UBUC) and 340 upper urinary tract UC (UTUC) patients. We observed that high SFRP2 expression in stroma but not in tumors is significantly linked to aggressive UC features, including high tumor stage and histological grade, positive nodal metastasis, the presence of vascular and perineural invasion, and high mitotic activity in UBUC and UTUC. Moreover, high stromal SFRP2 expression significantly and independently predicted worse clinical outcomes in UBUC and UTUC. Utilizing bioinformatic analysis, we further noticed that stromal SFRP2 may link epithelial-mesenchymal transition (EMT) to UC progression.

Conclusion: Collectively, these results imply that stromal SFRP2 may exert oncogenic function beyond its Wnt antagonistic ability, and stromal SFRP2 expression can provide prognostic and therapeutic implications for UC patients.

Keywords: SFRP2; bladder cancer; collagen; stroma; upper urinary tract cancer; urothelial carcinoma.

Figure 1

Expression profiles of the top 8 genes correlated with the Wnt signaling pathway in advanced/metastatic bladder tumors. The expression levels of upregulated and downregulated genes are marked in red and green, respectively. A comparative analysis (invasive vs. noninvasive and metastatic vs. nonmetastatic) was conducted under supervision. We identified SFRP2 as the most considerably upregulated gene related to the Wnt signaling pathway (GO: 0016055) among bladder cancer patients with advanced/metastatic disease.

Figure 2

High SFRP2 mRNA level is correlated with advanced stage disease, inferior overall survival, and CAF infiltration. (A) The correlations between the mRNA levels of SFRP2 and bladder cancer progression. (B) The impact of SFRP2 mRNA levels on overall survival in bladder cancer. These data were acquired from the GEPIA database. (C) The correlations among the mRNA levels of SFRP2, tumor purity, and CAF infiltration in bladder cancer. These data were estimated using the MCP-COUNTER and TIDE algorithms from the TIMER2.0 database. BLCA, bladder urothelial carcinoma.

Figure 3

High stromal SFRP2 immunoexpression was observed among bladder cancer patients with advanced stage disease. Immunohistochemistry was performed with an anti-SFRP2 antibody. SFRP2 immunoreactivity in stroma was gradually increased from (A) non-muscle-invasive to (B) muscle-invasive bladder cancer (200x, scale bar = 100 μm). Insect: 200x, scale bar = 25 μm.

Figure 4

Survival analysis. Kaplan–Meier curves show that high stromal SFRP2 immunoexpression conferred unfavorable prognostic effects on disease-specific survival and metastasis-free survival in (A–D) UTUC and (E, F) UBUC patients. Low tumoral SFRP2 expression was adversely prognostic only for metastasis-free survival in (G, H) UBUC patients. Also, high SFRP2 immunoexpression in stroma and low SFRP2 immunoexpression in tumors were significantly correlated with poor local recurrence-free survival in (I, J) UBUC patients.

Figure 5

The prominent GO terms enriched in SFRP2 upregulation. (A) The top 194 overlapping genes (Spearman’s correlation ≥ 0.6) co-upregulated with SFRP2 between UTUC and UBUC were examined utilizing the GO classification system according to (B) molecular functions, (C) cellular components, or (D) biological processes and were ranked by fold enrichment for functional annotation. To plot representative GO terms, an R script with ggplot2 package was used.

Figure 6

Gene coexpression network. A weighted network that connects key genes to each other was built using the GeneMANIA prediction server. Red and blue semicircles represent genes that belong to mononuclear cell proliferation and lymphocyte proliferation, respectively. Purple and grey lines indicate coregulation and pathways, correspondingly.